1. Field of the Invention
The present invention relates to gradient liquid transfer devices and methods for nano/micro liquid chromatographs, and more-particularly, to an improvement of liquid transfer performance in solvent mixing.
2. Prior Art
Recently, high performance liquid chromatographs have served as representative means for the separation and analysis of trace components. FIG. 4 shows an example structure of such a chromatograph. In a high performance liquid chromatograph 10, a solvent is transferred by a pump 14 from a mobile-phase solvent tank 12 through a passage, and an injector 16 introduces a trace sample into the solvent in the passage. The flow of the solvent leads the trace sample to a subsequent column 18. The trace sample is separated into components at a controlled temperature. The separated components are then detected by detection means 20, such as an absorbance detector. A computer 22 processes and analyzes A/D-converted detected signals, and also controls the conditions of the chromatograph.
Recently, a lower flow rate and a lower volume have been used to separate infinitesimal components at high resolution. Nano/micro liquid chromatographs which u se a flow rate of several tens of micro liters per minute to a nano liter per minute have been developed.
FIG. 5 shows a conventional liquid transfer system used for gradient elution in such a nano/micro liquid chromatograph. In the liquid transfer system 30, at a primary-solvent liquid transfer section 32, a primary solvent is replenished from a primary-solvent tank 34 to a metering pump 36, and then, a valve 38 is switched to transfer the primary solvent from the metering pump 36 through a primary-solvent passage 40 connected thereto.
At a secondary-solvent liquid transfer section 42, a secondary solvent is replenished from a secondary-solvent tank 44 to a metering pump 46, and then a valve 48 is switched to transfer the secondary solvent from the metering pump 46 through a secondary-solvent passage 50 connected thereto.
A three-way tee 52 (solvent mixing section) connected to the primary-solvent passage 40 and to the secondary-solvent passage 50 mixes the two solvents at a predetermined ratio. The mixed solvent is transferred to a subsequent separation system through a mixed-solvent passage 54. The mixing ratio of the two solvents is determined by the ratio of the flow rates specified by the metering pumps, and is controlled by a control section such as a computer. With the mixing ratio of the solvents being gradually changed, the mixed solvent is transferred to the subsequent separation system for gradient elution.
Due to the following two reasons, however, a considerable amount of the primary solvent enters the secondary-solvent passage 50 when only the primary solvent is transferred at a first stage.
First, since only a part (for example, from the pump 46 to the valve 48 in FIG. 5) of the passage 50 is initially filled with the secondary solvent, the primary solvent enters an empty part (for example, from the valve 48 to the three-way tee 52 in FIG. 5) of the passage 50, which has a considerable volume.
Secondly, the pressure of the liquid transfer system reaches, for example, as high as several tens of kilograms per square centimeters because of the resistance of the separation system, which is subsequent to the liquid transfer system. The primary solvent having a high pressure and entering the secondary-solvent passage 50 pushes the secondary solvent, which was previously stored at atmospheric pressure, to compress the secondary solvent. As a result, a further amount of the primary solvent enters the secondary-solvent passage. When water is used as the primary solvent and acetonitrile is used as the secondary solvent, acetonitrile is compressed due to the pressure of the water, and a considerable amount of volume contraction occurs. Since nano/micro liquid chromatographs have very small passage diameters and very small passage volumes, this solvent contraction also largely affects the entry of the primary solvent into the secondary-solvent passage.
When a considerable amount of the primary solvent enters the secondary-solvent passage in this way, even if the operations of the metering pumps are controlled such that the transfer of the primary solvent only is switched to the transfer of the mixed solvent at a predetermined time, actual switching is performed at a time later than the predetermined time, as shown in the graph of FIG. 6. In other words, it takes time for the pump 46 to push back the primary solvent which has entered the secondary-solvent passage 50, the secondary solvent cannot be mixed with the primary solvent during this time; and a time delay thus occurs. Therefore, the mixed solvent may be transferred to the separation column with a delay, so that separation is performed late; or analysis of the measurement results may be inaccurate, which is a problem.